Effect of ammonia on the glutamate dehydrogenase catalyzed oxidative deamination of L-glutamate: production of an ammonia-containing intermediate in the "burst" phase

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Abstract
The effects of ammonium acetate on the transient burst phase of the oxidation of L-glutamate by glutamate dehydrogenase were studied. Two measurable changes are observed in the burst phase as ammonium acetate concentration is increased: an increase in the apparent 1st-order rate constant, kapp, and a decrease in the amplitude of the absorbance change measured at 320 nm. The increase in kapp shows a hyperbolic dependence on ammonium acetate concentration and is independent of glutamate concentration. The existence of an intermediate immediately following H transfer was demonstrated. The intermediate contains enzyme, reduced coenzyme, ammonia and .alpha.-ketoglutarate moieties and is in equilibrium with the known complex consisting of enzyme, reduced coenzyme and .alpha.-ketoglutarate. At high concentrations of ammonium acetate, the equilibrium favors the ammonia containing complex.